Manual dako imunohistoquimica

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PATHOLOGY

Education Guide

Immunohistochemical Staining Methods
Fourth Edition

Dako provides cancer diagnostic
products for leading reference
laboratories, hospitals and other
clinical and research settings.
Our instrumentation portfolio is
complemented by a full line of
antibodies, pharmDx™ assays,
detection systems and ancillaries.
Consider Dako for all your
laboratory needs. PATHOLOGY

Education Guide

Immunohistochemical Staining Methods
Fourth Edition

Dako provides cancer diagnostic
products for leading reference
laboratories, hospitals and other
clinical and research settings.
Our instrumentation portfolio is
complemented by a full line of
antibodies, pharmDx™ assays,
detection systems and ancillaries.
Consider Dako for all your
laboratoryneeds.

Immunohistochemical Staining Methods,
Fourth Edition
Editor
Marc Key, Ph.D.
Key Bomedcal Servces ° Oja, CA, USA

Contributors
Karen Atwood, B.S. MT (ASCP) CLS
Dako ° Carpntera, CA, USA

Kirsten Bisgaard, B.S.
Dako ° Glostrup, Denmark

Kenneth J. Bloom, M.D.
Clarent ° Also Vejo, CA, USA

Thomas Boenisch, M.S.
Dako ° Carpntera, CA, USA

Nanna K. Christensen,M.S., Ph.D.
Dako ° Glostrup, Denmark

A.J. Farmilo, Ph.D.
Dako ° Mssssauga, Ontaro, Canada

Richard Harvey, Ph.D.
Unversty of New Mexco School of Medcne ° Albuquerque, NM, USA

Jim Hudson, Ph.D.
Dako ° Carpntera, CA, USA

Mehrdad Nadji, M.D.
Unversty of Mam School of Medcne ° Mam, FL, USA

W. Roy Overton, Ph.D.
GCAT Inc. ° Fort Collns, CO, USA

Gale E. Pace, B.S.MT (ASCP) I
Dako ° Carpntera CA, USA

Ole Feldballe Rasmussen, M.S., Ph.D.
Dako ° Glostrup, Denmark

Andreas Schønau, M.S., EBA
Dako ° Glostrup, Denmark

Helle Grann Wendelboe, M.S.
Dako ° Glostrup, Denmark

Lars Winther, M.S., Ph.D.
Dako ° Glostrup, Denmark

Ron Zeheb, Ph.D.
The Lahey Clnc ° Burlngton, MA, USA

© Copyright 2006 Dako, Carpinteria, California. All rightsreserved. No part of this book may be
reproduced, copied or transmitted without written permission. US $50



Immunohstochemcal Stanng Methods

Legend for Schematics

primary
antibody

HRP
AP
enzyme enzyme



antibody

secondary
antibody

biotin streptavidin biotinyl
label
t yramide

antibody

fluorescein
label

F(ab1)2
secondary
antibody

DBA

Fast Redtissue
antigen

polymer

Immunohstochemcal Stanng Methods

Table of Contents
Preface VI
ParT I: Theory
Chapter 1 ° antibodies
Thomas Boenisch
Introduction 1, Immunoglobulins 1, IgG 2, IgM 3, Polyclonal Antibodies 5,
Monoclonal Antibodies 6, Antibody Affinity 7, Antibody Cross-Reactivity 9,
Antibody Reaction Rates 10, Antibody Stability 10, Handling of Antibodies 12
Chapter 2 °Basic Immunochemistry
Thomas Boenisch
Introduction 15, Antibody Titer 15, Antibody Dilution 15, Antibody Incubation 17
Chapter 3 ° Basic Enzymology
Thomas Boenisch
Introduction 19, Enzymes 19, Substrates and Chromogens 22, Suggested
Procedures for Substrate-Cromogen Reagents 23
ParT II: Processing
Chapter 4 ° Fixation and Processing
A.J. Farmilo
Introduction 27, Fixation 27, TissueHandling 29, Specialized Tissue
Preparations 31
Chapter 5 ° Molecular-Friendly Tissue Processing
Mehrdad Nadji
Why Molecular Pathology 35, Complete Molecular-Friendly Histology
Platforms 35, Fixation 35, Processing 36, Validation 37, Summary 37,
Conclusion 39
Chapter 6 ° antigen retrieval
Marc Key and Thomas Boenisch
Introduction 41, Short History of Antigen Retrieval 41, Principle andTechnique 42,
Mechanism of Action 43, Cytology 43, Target Retrieval for In Situ Hybridization 44,
Antigen Retrieval and Its Use in Double Staining 44, Conclusion 44



Immunohstochemcal Stanng Methods

ParT III: Methodologies
Chapter 7 ° Immunohistochemistry Staining Methods
Marc Key
Introduction 47, Immunohistochemistry 47, Avidin-Biotin Immunohistochemistry
48, Polymer-Based...
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